Multiple analyte assay device

ABSTRACT

A test device and associated method of use composed of a housing ( 100 ) defined by base ( 101 ) and cover ( 110 ). The base ( 101 ) has a closed end ( 104 ) and an open end ( 106 ). Slots ( 102   a,    102   b , etc.) are formed within the base ( 101 ) and are separated by rails ( 103   a,    103   b , etc.) where test strips ( 105   a,    105   b , etc.) are inserted.

FIELD OF THE INVENTION

The present invention relates to methods and devices for assayingbiological fluid samples. More particularly, the invention relates tomethods and devices for detecting analytes, such as drugs, in urine.

HISTORY OF THE RELATED ART

In their most simple form, chromatographic analyte test strips permit anassay to be performed in a single step (application of an analyte sampleto the device) to produce visually observible assay results (such asthose indicated by colored bars on the test strip). However, a commonlimitation of such test strips is that they can only be used to detect asingle analyte, requiring that serial assay procedures be performed todetect additional analytes (for example, to test a sample for thepresence of a panel of narcotics). Multiple dipping steps, such as arecommonly used when multiple dipstick assays are separately performed,present not only a possible loss of sensitivity of the assay (throughreagent mixing or loss of reagent solutions), but also an esthetic andhygienic problem for the analyst. Repetitive performance of assayprocedures is also tedious, which increases the risk that assays will beperformed improperly or the results misinterpreted.

SUMMARY OF THE INVENTION

The present invention provides an assay device, device for separating afluid analyte sample for use in multiple assay procedures and methodsfor performing multiple analyte assays. In one embodiment of the assaydevice, the assay device is a dipstick having multiple analyte teststrips, each of which includes a test zone and a control zone. The teststrips are enclosed in a housing having an open side through which anend of each test strip protrudes to form a sample loading zone. Aprotective cap is provided to seal the protruding ends of the teststrips from exposure while not in use. Each test strip is separated fromthe next within the housing by a raised spacer. The portion of thehousing which overlies the test and control zones is transparent topermit visually observible results shown in each zone to be viewed.

In cassette form, the assay device has the same structure describedabove, but the protruding test strips are inserted into a cap which hasa sample port for application of sample to the test strips. The cap isretained on the assay device by a close fit over the device housing.

Each test strip provides binders and assay reagents for detection of adifferent analyte in the sample fluid. In a particularly preferredembodiment of the assay device, the housing may be opened to permitsubstitution of different test strips to allow each device to becustomized for detection of specific analytes of interest. Assay sampleintegrity determinants consisting of test strips which allow measurementof parameters such as specific gravity and pH may also be included ineach device.

The invention also provides a separator device for dividing a fluidassay sample into portions for use in multiple assays without need forcontact between the assay operator and the fluid sample. This latterfeature of the device increases operator safety and avoids inadvertentcontamination of the assay sample. The separator device may be used toseparate any fluid assay sample, but is especially useful in assayingsamples for the presence of narcotics, where a positive result on firsttesting of the sample may necessarily be followed by additional testingof the sample to confirm the result and the identity of the detectednarcotic. To this end, the separator device is adapted particularly wellto use with the assay device of this invention.

The assay device of the invention makes specimen analysis easier becausean analyte sample need only to be applied once to the assay device fortesting. In addition, the replaceable nature of the analyte test stripsallows the analyst to customize the array of assays to the testingsituation. Because the customization can be performed before adding thetest sample (e.g., urine), fewer manipulations with the analyte sampleare needed to obtain the desired information. In addition, use of theseparator device permits further testing of the sample to be performedwithout risk of adultering the sample in a preliminary assay performedaccording to the invention.

The invention also provides a method for assaying one or more analytesof interest. The protruding ends of the device are dipped into a fluidanalyte sample. Binding of an analyte present in the sample with one ormore specific ligands causes formation of specific visual pattern in thetest and control zones indicative of the test result. The assay resultsperformed according to the invention may be read visually without use ofseparate measuring equipment. Thus, performance of assays according tothe invention requires only that the user introduce the requisite amountof test sample into the device of the invention, then observe any colorchanges which appear shortly thereafter in a detection zone of ananalyte strip. The method of the invention is especially useful forscreening fluid analyte samples (e.g., urine) for the presence orabsence of drugs of abuse.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is an exploded view of a dipstick assay device of the invention.

FIG. 2 is a top view of a dipstick assay device of the invention.

FIG. 3A is a top view of the upper half sample port cap of a cassetteassay device of the invention, and FIG. 3B is a top view of the lowerhalf, base portion of the sample port cap, while FIG. 3C is a side,cut-away view of the intact cap with test strips in place therein.

FIG. 4 is a top view of the separator device of the invention.

FIG. 5 is a cross-sectional view of the separator device taken alongline A-A at cut-away point B-B of FIG. 1.

FIG. 6 is a lateral view of the separator device within a specimencollection cup.

Like numerals refer to like elements in the drawings.

DETAILED DESCRIPTION OF INVENTION A. Definitions

For ease of understanding, the following definitions will applythroughout this description:

1. The term “antigen” as used herein refers to any analyte which iscapable of binding antibodies. Antigens may comprise, withoutlimitation, chemical compounds, polypeptides, carbohydrates, nucleicacids, lipids, and the like, including viral particles, viral subunits,bacterial and parasite surface antigens, and host proteins that may bediagnostic of the subject's condition.

2. A “binder” refers to a ligand for the analyte as in the format of asandwich assay, or a ligand for both the analyte and the tracer as inthe format of a competitive assay. A binder can be chosen from a groupof molecules or compounds capable of binding the analyte, such as anantigen to the antibody analyte, or an antibody to the antigen analyte.

3. A “test zone” refers to an area in which a binder or the analyte isattached, movably or immovably, to the test strip portion of an assaydevice.

4. A “tracer” refers to a ligand for the analyte or the binder labeledwith a detectable label, preferably a visually readable particulatelabel, such as colloidal gold, latex and liposomes including dye, carbonblack, and the like.

5. A “sample loading zone” refers to an area of a test strip on which afluid analyte sample is applied for migration to the test zone.

6. A “test strip” of the invention consists of, collectively, all of thezone supporting membranes and any filters of the assay device.

7. A “fluid analyte sample” can be any fluid suspected of containinganalyte of interest for which a particular assay is specific. Testsample may represent any body fluid, including urine, blood, sweat,lymph, intraperitoneal fluid, crude tissue extract or homogenate,derived from a fetus, neonate, juvenile or adult subject; anon-biological fluid such as water from some ecological niche, e.g., ariver or a lake; or a solution used in a laboratory.

8. A “label” is a molecule or compound which directly or indirectlymediates the formation of a signal (such as a color change) which isused in assay to indicate the presence, absence or concentration rangeof analyte of interest in a test sample. Labels may include enzymes,fluorescers, liposomes, erythrocyte ghosts, polymer microcapsules, colorpolymer particles (latex), and preferably includes sols ofmetal-containing compounds. A wide variety of patents and patentapplications provide an extensive literature of different techniques forproducing detectible signals in immunoassays. The following list ofUnited States patents is merely illustrative of the type of label whichcan find application in this invention: U.S. Pat. Nos. 3,646,346discloses radioactive label; 3,654,090, 3,791,932, and 3,817,838disclose enzyme labels; 3,996,345 discloses fluorescer-quencher labels;4,062,733 discloses radioactive label; 4,067,959 discloses fluorescer orenzyme label; 4,104,099 discloses chemiluminescent label; and 4,160,645discloses non-enzymatic catalyst label. U.S. Pat. No. 3,966,879discloses an electrophoretic technique employing an antibody zone andU.S. Pat. No. 4,120,945 discloses a radioimmune assay (RIA) wherelabeled analyte is initially bound to a solid support through antibody.U.S. Pat. No. 4,233,402 discloses enzyme pair labels; U.S. Pat. No.4,720,450 discloses chemically induced fluorescent labels; and U.S. Pat.No. 4,287,300 discloses enzyme anionic charge labels.

Labels can also be metal-containing sols; i.e., metal or metal compoundssuch as metal oxides, metal hydroxides, metal salts, metals ormetal-containing compounds mixed with polymers or coated onto polymernuclei. These metal labels may include dry forms of any of theabove-named metal or metal compound sols, and preferably includescolloidal gold in dry form.

9. A “complex” means (depending on the context) any multimolecularcomplex formed by analyte and one or more ligands, or by labeled ligandand immobilized ligand. In a sandwich-type immunoassay, e.g., thefollowing complexes occur: analyte/labeled ligand duplex first producedin the assay (first complex) and analyte/labeled ligand/immobilizedligand triplex formed second in the assay (second complex).

10. “Fluid communication” refers to structures which are in contactwith, but not necessarily affixed to, one another.

11. “Assay” refers to several different types of assay formats in whichan analyte of interest can be detected using an assay test strip. Forexample, in a sandwich-type immunoassay, analytes of interest in theanalyte sample, when present, bind a labeled tracer movably incorporatedin the test strip (consisting of a porous membrane) at the tracer zoneto form a first complex. The tracer is a molecule which binds theanalyte of interest and is conjugated to a label, preferably a metallabel, and most preferably colloidal gold.

A second immobilized ligand corresponding to the analyte of interest iscoupled to the test strip at the test zone. First complex and unboundlabeled ligand mix with the test sample and be carried along therewithby capillary action (wicking) through the test zone. Analyte samplepasses through the test strip bringing the first complexes, if any, intocontact with the unlabeled ligand immobilized in the test zone to form asecond complex of labeled ligand-analyte-immobilized ligand. The firstimmobilized ligand is immobilized in the test zone by means known in theart, including covalent bonding or attachment to an insolubleprotein-coated surface (see, e.g., U.S. Pat. Nos. 4,200,690 and5,075,078). When the second complex is formed, a visible color patternappears in the test zone. Labeled ligand not bound to analyte in thetest sample continue migration by wicking into the control zone tocontact the ligand immobilized there. The labeled ligand can bind theimmobilized ligand in the control zone to form a third complex, and thusbe captured in the control zone.

Within the scope of this invention, the labeled ligand forming thecomplex in the control zone may be the same as the tracer forming thefirst and second complexes, or it may be a different labeled ligand. Theligand immobilized in the control zone should have specific affinity forthe labeled ligand intended to form the third complex. Formation of thethird complex is indicated by a visible pattern in the control zone.

Besides sandwich immunoassay method, other assay methods may beimplemented in the devices of the invention. These methods may includecompetition and inhibition assays. In a competition assay, the analyteand tracer have similar affinity properties and compete for binding withimmobilized ligand. Thus, in absence of analyte, the pattern (e.g.,band) in the test zone is of maximum intensity. When present, theanalyte binds to immobilized ligand to prevent the tracer from gettingcaptured in the test zone. Thus, the intensity of the test band isreduced, depending on the concentration of analyte in the test sample.

In an inhibition assay, the analyte and immobilized ligand in the testzone each have affinity for the tracer. In the absence of analyte in theanalyte sample, the tracer is captured by immobilized ligand, and avisible pattern forms in the test zone. When present, the analyte bindsthe tracer, thereby preventing it from binding to the immobilized ligandin the test zone. The resulting intensity of the test band is reduceddepending on the concentration of analyte in the test sample.

B. Dipstick Assay Device

Turning to FIG. 1, a dipstick form of the assay device is shown inexploded view. The device consists of a housing 100, which is defined bybase 101 and cover 110. Base 101 can be constructed of any sterilizablematerial, such as a nonporous plastic (e.g., the commercially availableplastic “ABS” supplied by the Monsanto Company of St. Louis, Mo.). Base101 having a closed end 104 and an open end 106, slots 102A, 102B, 102C,102D and 102E separated by rails 103A, 103B, 103C and 103D for insertionof test strips 105A, 105B, 105C, 105D and 105E. A particular advantageof this embodiment of the assay device is its customizability in thattest strips specific for different analytes of interest to the user maybe inserted into base 101 and that the number of test strips employedmay vary (e.g., base 101 may have any number of slots from two upward toaccommodate as many test strips as the user may desire).

Referring to FIG. 2, when inserted into slots 102A, 102B, 102C, 102D and102E, the test strips extend out of base 101 beyond open end 106. Thelength to which the test strips protrude from base 101 must besufficient to allow the test strips to contact a fluid analyte sample,preferably by immersion, and most preferably without allowing the fluidto contact housing 100. The test strips are conventional in form;therefore, because those of ordinary skill in the art will be abundantlyfamiliar with the design of such test strips, they will not be describedin detail here. However, each test strip will have a test zone 112 forbinding of analyte (to indicate a positive test result for the presenceof analyte in the analyte sample) and a control zone 113 for binding oftracer (to indicate correct operation of the assay). Preferably, thetest zones and control zones of each test strip lie in the same locationon each test strip so each can be viewed in side-by-side fashion.

Each test strip is typically constructed of a porous membrane which issubstantially inert with respect to the analyte and must be porous orabsorbent relative to the analyte sample to be tested, e.g., urine. Thesubstance can be either bibulous matrices or nonbibulous matrices thatare insoluble in, and maintain their structural integrity when exposedto aqueous solutions or physiological fluids. Bibulous matrices that canbe useful for the devices of the present invention include but are notlimited to, paper, sponge materials, cellulose, hydrophilic inorganicpowders, wood, synthetic resin fleeces, woven and nonwoven fabrics andlike materials. Nonlimiting examples of nonbibulous matrices includeglass fiber, permeable polymer films and preformed or microporousmembranes. The absorbent material is preferably absorbent paper. Theabsorbent material can be affixed by a double sided adhesive (e.g., twosided adhesive tape) to a solid moisture impervious support. Thissupport can be constructed from, for example, hydrophobic plastic,cellulose acetate, polyethylene, terephthalate, polycarbonate, orpolystyrene.

The tracer is prepared according to the means known in the art. Forpurposes of producing a clearly visible reaction, labels ofmetal-containing sols are preferred, with labels of colloidal gold orselenium being most preferred. An example of a suitable product iscolloidal gold available from Janssen Life Sciences Products. Thesecolloidal metals produce distinctive visual patterns without addition offurther reagents; however, fluorescers (such as fluorescein) and enzymes(such as those identified in U.S. Pat. No. 4,275,149), may also be used.

Selections and choices for test binders (e.g., immobilized antigens,antibodies and other test and control binders), as well as suitablemeans for their attachment to porous test strip membranes, arewell-known to those of ordinary skill in the art and will not be statedin detail here. To maximize contact of test sample with the tracer andall test binders, the area occupied by each reagent on the test strippreferably extends from one side of the membrane to the other.

For further review concerning test strip construction, includingselection and preparation of test reagents, the following referencesprovide a representative sample of test strip designs known in the art:U.S. Pat. No. 5,384,264 (commonly owned); U.S. Pat. No. 4,491,645; U.S.Pat. No. 4,943,522; U.S. Pat. No. 5,252,496; U.S. Pat. No. 5,714,389 andU.S. Pat. No. 5,602,040, the disclosures of which are incorporated forpurposes of reference.

Test strips 105A, 105B, 105C, 105D and 105E may be secured within slots102A, 102B, 102C, 102D and 102E by adhesion to the floor of each slot;however, the placement of cover 110 onto base 101 is sufficient toretain the test strips within the base slots. To this end, cover 110 isconveniently constructed of an opaque tape having at least onetransparent window 111 formed therein for viewing of test results alongtest zone 112 and control zone 113. To secure cover 110 onto base 101,as well as to secure test strips 105A, 105B, 105C, 105D and 105E withinslots 102A, 102B, 102C, 102D and 102E, cover 110 is pressed into placeto form an adhesive attachment between cover 110 and the upper edges ofrails 103A, 103B, 103C, and 103D. To provide additional surface area foradhesion of cover 110 to base 101, bar 107 separates closed end wall 104of base 101 from rails 103A, 103B, 103C, and 103D.

Conveniently, cover 110 is also provided with transparent windows 115A,115B, 115C, 115D and 115E through which labels on test strips 102A,102B, 102C, 102D and 102E can be viewed. The labels (not shown) may beprinted with information of use in performing the assay, such as theidentity of analyte detectible with each test strip.

In certain instances, it may be desirable to store the assay deviceafter test results are obtained for later viewing. To that end, afive-sided cap 120 is provided for insertion over open end 106 of base101 (with cover 110 in place) to protect the protruding ends of teststrips 102A, 102B, 102C, 102D and 1102E from contact with othermaterials, from dessication and from contact with the assay operator.Cap 120 is easily secured onto the assay device by a close fit, such asa friction fit or snap-fit.

C. Cassette Assay Device

In some instances (e.g., where the analyte sample is believed to containpathogenic organisms) it is desirable to protect the assay operator fromcontact with analyte sample after its application to an assay device. Tothis end, the dipstick assay device may be conveniently modified for usein closed cassette form.

More specifically, cap 220 (FIGS. 3A and 3B) is adapted to convert thedipstick assay device into a cassette. Cap 220 is similar in design tocap 120 (FIGS. 1 and 2), except that sample application slot 221 isformed therein to permit analyte sample to be applied to test strips202A, 202B, 202C, 202D and 202E dropwise; e.g., by pipetting the samplethrough slot 221 (in FIG. 3C, only strip 202A is visible from the sideview and the device housing is not shown). To avoid sample overflow, areservoir 222 may be provided in the inner floor 223 of cap 220 by, forexample, providing raised bar 226 on floor 223 (in FIGS. 3A and 3B,floor 223 is shown as if split from roof 225 of the cap only for thepurpose of permitting reservoir 222 to be viewed in the drawing). Adownwardly protruding bar 224 is provided from the inner surface of roof225 of cap 220 to depress the test strips into reservoir 222 so eachtest strip has equal access to the analyte sample. After performance ofthe assay, cap 220 remains in place on the assay device to protect theprotruding ends of the test strips from contact with other materials,from dessication and from contact with the assay operator.

D. Separator Device for Division of Analyte Sample

If a positive result is obtained from use of the assay device of theinvention, it is usually necessary to further characterize the detectedcompound to better enable its identification; e.g., by massspectrometry. However, it is rarely practical to ask that more than oneassay sample be obtained from a subject. As such, any assay sample whichis obtained must be divided into portions of sufficient volume forrepeated testing, for example by pouring the original sample intoseparate specimen containers (at the risk of operator contamination andsample loss). Even where the sample is only to be assayed once, thetendency of subjects to provide abundant urine samples poses a differentproblem in that too much sample can saturate a test strip and overwhelmthe assay reagents. Again, division of the sample is required.

The separator device of the invention provides simple means for dividinga sample while protecting the sample from contamination and the operatorfrom the sample. To these ends, the separator device consists of a ringwhich is just smaller in diameter than the inner diameter of the openend of a specimen collection cup so, when pressed inside of the open endof the cup, the ring will remain seated there. A collection chamber (forexample, a “V” shaped well) extends across the ring and is attachedthereto so the ends of the collection chamber are closed by the innerwalls of the ring.

In use, an assay fluid is placed within the specimen collection cup to alevel below the point where the separator device will be seated. Theassay operator presses the separator device into place and seals thespecimen collection cup with a cap. The operator inverts the specimencollection cup several times so fluid pours into the collection chamberof the separator device. The balance of the fluid assay remains belowthe level of the separator device and is therefore protected fromcontact with reagents or other material placed therein. A test strip(such as the assay device of the invention) is placed into contact withthe portion of the assay sample contained within the collection chamberof the separator device; e.g., by dipping an end of the test strip intothe collection chamber. After the assay sample is loaded from thecollection chamber onto the test strip, the latter is removed and theseparator device is carefully lifted from the specimen collection cupfor disposal.

Use of the separator device provides the assay operator with a volume ofassay sample fluid which is sufficiently limited to avoid saturation ofthe test strip. For example, where the assay device utilized is thedevice of this invention, the collection chamber is of a depth andlength sufficiently limited so the maximum fluid level achievable in thecollection chamber is lower than the level of the assay device housing.The uncontaminated balance of the assay sample still in the specimencollection cup is available for further testing; e.g., for massspectrometry to determine the identity of any compounds detected in theinitial assay of the portion of the sample separated in the separatordevice. Conveniently, the separator device may be provided in the formof a kit, including the separator device, a sterilized specimencollection cup with cap and forceps for removal of the separator devicefrom the cup after performance of the assay. Such a kit may also beprovided with the assay device of the invention.

An example of a separator device is shown in FIGS. 4 through 6. Althoughthe separator device shown is in the shape of a ring (to correspond tothe common cup-like shape of urine collection cups), those of ordinaryskill in the art will recognize that the separator device may be of anyshape which conforms to a specimen collection container having at leastone open end into which the separator device may be seated.

Referring to FIG. 4, a top view of separator device 400 is provided.Ring 401 has an OD of slightly less than the ID of the specimencollection cup into which the separator device is to be placed.Collection chamber 402 spans ring 401 and is defined by walls 403 and404.

Looking through separator device 400 along line A-A from cut-away pointB-B (FIG. 5), it is seen that walls 403 and 404 meet at point 405 toform a V-shaped well. Those of ordinary skill in the art will recognizethat collection chamber 402 may take the form of a half-circular,squared or other shaped well, but a V-shape is a convenient well form tomanufacture. Collection chamber 402 is closed at both ends of the wellby inner surface 406 of ring 401. Also, it will be appreciated thatseparator device 400 may be provided with more than one collectionchamber.

FIG. 6 shows separator device 400 in place within specimen collectioncup 410. It can be seen from FIG. 6 that the height of ring 401 isrestricted so neither it nor collection chamber 402 extend above thelevel of the open mouth 411 of cup 410 (to avoid interfering withclosure of cup 410 by its cap [not shown]). Assay sample fluid 412remains below separator device 400 in cup 410.

Separator device 400 may be constructed of any sterilizable materialwhich is acceptable for use with fluid assay samples, the identity ofwhich will be known to those of ordinary skill in the art (e.g.,plastics such as polycarbonate and glass). Preferably, the material willbe non-porous and hydrophobic.

E. Methods for Use of the Assay Devices

The method of the invention may be used to detect any analyte present influid sample. The invention is especially useful for detection ofmonoepitopic and polyepitopic antigens and antibodies associated withpathologies, as well as physiological compounds and drugs.

The assay devices of the invention are particularly well suited for usein drug screening assays and for diagnostic testing of organisms. In theformer respect, a five drug panel of assay tests is recommended by theNational Institute on Drug Abuse (NIDA), which includes tests fortetrahydrocannabinol and other marijuana metabolites, cocainemetabolites, opiate metabolites, phencyclidine (PCP, Angel Dust), andamphetamines. For a more extensive substance abuse testing panel, thechoice of analytes tested can include marijuana metabolites;tetrahydrocannabinol and other marijuana metabolites, cocainemetabolites, opiate metabolites, phencyclidine (PCP, Angel Dust),amphetamines, barbiturates, benzodiazepines, methaqualone, andpropoxyphene. The analyte test strips for drug tests preferably have thesensitivity equal to the cutoffs recommended by Substance Abuse MentalHealth Service Administration (SAMSHA) and NIDA, which most employersuse. Binders and reagents for use in constructing test strips for use indetecting drugs of abuse are well-known in the art and will not bedescribed in detail here; however, representative sources of suchmaterials are described in the Examples below.

Subjects undergoing drug tests are often creative in their attempts toadulterate the analyte samples to evade detection of drugs of abuselikely to be present in the sample. To minimize the effects of suchevasion efforts on results obtainable with the assay devices of theinvention, test strips may be provided in the devices which indicate theintegrity and condition of the analyte sample. For example, test stripsmay be provided to simultaneously assay the analyte sample for pH,osmolality (the total concentration of solutes in urine, expressed asmOsm/kg and measured as a function of fluid specific gravity), or thepresence of albumin.

In test strips for pH, the strip is impregnated with various dyes thatrespond with different color changes to a pH in the range of 5 to 9.Depending on the acid-base status, urinary pH may range from as low as4.5 to as high as 8.0. Although this test is done routinely, it neitheridentifies nor excludes patients with urinary system disease. The testcan, however, indicate that the condition of the urine sample hasdeteriorated.

To test the specific gravity (which is directly proportional to urineosmolality) of an analyte test fluid, analyte test strips are availablethat measure specific gravity in approximations. For example, U.S. Pat.No. 4,318,709, to Falb et al., issued Mar. 9, 1982, provides a testmeans for determining the ionic strength or specific gravity of anaqueous test sample, the test means comprising a weakly acidic or weaklybasic polyelectrolyte which is at least partially neutralized, and anindicator means capable of producing a detectable response to ionexchange between the polyelectrolyte and the sample. The test device isa carrier matrix incorporated with the test means, and the method forits use involves contacting an aqueous test sample with the device andobserving a detectable response. The disclosure of the '709 patent isincorporated for reference herein.

Normal urine osmolality varies between 50 and 1200 mOsm/kg (specificgravity between 1.002 and 1.035). Any urine having a specific gravityover 1.035 is either contaminated, contains very high levels of glucose,or the patient may have recently received high density radiopaque dyesintravenously for radiographic studies or low molecular weight dextransolutions.

Commercially available analyte test strips also permit simple and rapidtesting for protein. Methods based on dye binding techniques have provenespecially useful because dye binding methods are readily automated andprovide reproducible and accurate results. In general, dye bindingtechniques use pH indicator dyes that are capable of interacting with aprotein, such as albumin, and that are capable of changing color uponinteraction with a protein absent any change in pH. When a pH indicatordye interacts with, or binds to, a protein, the apparent pKa (aciddissociation constant) of the indicator dye is altered and the dyeundergoes a color transition, producing the so-called “protein-error”phenomenon. In methods utilizing the dye binding technique, anappropriate buffer maintains the pH indicator even at a constant pH toprevent a color transition of the pH indicator dye due to a substantialshift in pH. Due to the “protein-error” phenomena, upon interaction withthe protein, the pH indicator dyes undergoes a color transition that isidentical to the color change arising because of a change in the pH.Examples of pH indicator dyes used in the dry phase assay of proteinsthat are capable of interacting with or binding to proteins andexhibiting “protein-error” color transitions include tetrabromophenolblue and tetrachlorophenol-3,4,5,6-tetrabromosulfophthalein. Simple,accurate and inexpensive protein detection assays have been developedfor the detection or measurement of protein in urine and serum (See,e.g., U.S. Pat. No. 5,096,833 to Lau et al., incorporated herein forreference).

The method of the invention is performed by applying analyte sample totest strips by immersion (dipstick forms of the device) or by applyingthe sample dropwise through slot 221 in cap 220 (FIG. 3; representingcassette forms of the device). After waiting a predetermined time, suchas from about 15 seconds to about 60 seconds, test results are viewedthrough window 111 or 211 (FIGS. 1 and 2), either visually or by aninstrument. A color change in test zone 112 or 212 (FIGS. 1 and 2)indicates the presence or concentration of analyte in the sample. Whenno band appears in test zones, or if the control band is neitherdistinct nor fully formed, the assay is regarded as incompetent toindicate the presence or absence of analyte in the test samples and maybe performed again. In addition, the assay can be made quantitative byemploying spectrophotometric or calorimetric techniques, as opposed tovisual techniques, in order to more reliably and more accurately measurethe degree of color transition, and therefore more accurately measurethe concentration of analyte in the test sample.

F. Kits

The invention provides a kit useful for the detection of analytes ofinterest, having a carrier compartmentalized to receive one or morecontainers holding the multianalyte assay device of the invention orparts thereof. Preferably, the multianalyte assay device is part of akit which may be composed of the device, instructions for its use, asample collection cup, a capillary device for measuring test sample, apipette for introduction of sample to the device, and a desiccantpacket.

Desiccant provides low humidity conditions necessary for preservation ofreagents during the shelf life of the device. Alternatively, a desiccanttablet or a desiccant packet may be included in an air-tight protectivepouch with the device. Instructions for use of the multianalyte assaydevice may be printed onto the cover or onto the packaging of themultianalyte assay device or may be printed in literature to be packagedwith the multianalyte assay device. The kit may additionally include anattached temperature strip, lids for the specimen cup, and theliterature. Components of such a kit for use in performing an assayprocedure (e.g., excluding printed instructions) are preferably to besealed in one or more air-tight packages, such as foil packets.

The following examples are provided to illustrate a use for theinvention and do not limit its scope. Unless otherwise noted, all termsand abbreviations used in the examples are standard in the art.

EXAMPLE 1 Assay for Six Drugs of Abuse

Six chromatographic strips for detecting drugs of abuse(methamphetamine, opiates/morphine, marijuana/tetrahydrocannabinol,amphetamine, cocaine/benzoylecgonine, benzodiazepine) each of a size of5 mm×73 mm were placed in slots of the device of the invention as shownin FIG. 1. Each strip consisted of a colloidal gold-labeled antibody(specific to the target drug) incorporated into the upstream end of thestrip (tracer zone) in the middle of a 30 mm fiberglass matrix, and anantigen-BSA binder immobilized in the center (binder zone) of a 22 mmnitrocellulose membrane lying downstream of, and in fluid communicationwith, the fiberglass matrix (wherein the antigen is either the drug ofinterest or an analog thereof having the same immunogenicity).Downstream to the nitrocellulose membrane was a 26 mm long filter paper.The matrix, membrane and filter paper were attached on a vinyl sheet soeach was in fluid communication, by overlapping 2 mm of each of theirends.

15 drops (0.7 ml each) of analyte sample (human urine) were applied tothe sample port. Results were read after 10 minutes. The presence orabsence of a pink-rose color band in the binder zone indicated negativeor positive results for the presence of each drug of interest in theanalyte sample.

For comparison, additional aliquots of the analyte samples wereseparately tested for the presence of the same drugs of abuse by acommercial assay (Syva EMIT EIA II). The second panel of test resultscorrelated with the results obtained according to the invention.

Although the invention has been described with reference to thepresently preferred embodiment, it should be understood that variousmodifications can be made without departing from the spirit of theinvention. Accordingly, the invention is limited only by the followingclaims.

1. A device for assaying a fluid for the presence or absence ofdifferent analytes comprising: (A) a base having adjacent slots thereinof sufficient length for insertion of a test strip therein, wherein eachslot is defined by (a) a floor, (b) raised walls depending upwardly fromthe floor to separate each adjacent slot from the next, and (C) at leastone open end; (B) a multiplicity of test strips having an upstream and adownstream end, wherein a single test strip is inserted into each slotof the base so the upstream end of each test strip protrudes out of theopen end of each slot, and wherein each test strip has a test zone and acontrol zone therein, and each test zone contains a binder specific froa different analyte; (C) a cover attached to the upwardmost surface ofeach raised wall of the slots of the base and extending to the open endof said base, wherein the cover retains the test strips within the slotsand has a first transparent window formed therein through which the testzone and the control zone of each of the test strips can be viewed; (D)said base and said cover forming the housing for said test stripsextending beyond that housing; and (E) a removable cap over said baseand cover enclosing the protruding ends of the test strips extendingbeyond the housing, the cap also containing a sample port and opposingsaid port a base having a raised bar therein which defines a fluidreservoir beneath the sample port whereby the fluid analyte sample maybe applied to the protruding ends of the test strips when in place overthe strips.
 2. The device according to claim 1 further comprising asecond transparent window formed within the cover through which the teststrips can be viewed.
 3. The device according to claim 1 furthercomprising a multiplicity of test strips inserted into each slot of thebase, wherein each test strip has a test zone therein and each test zonecontains a binder specific for a different analyte.
 4. The deviceaccording to claim 3 wherein each binder is specific for a differentdrug of abuse.
 5. The device according to claim 2 wherein each test zoneis visible through the first transparent window of the cover.
 6. Thedevice according claim 2 each test strip further comprises a labeldownstream of the test zone, which label identifies the analyte forwhich the binder is specific.
 7. The device according to claim 6,wherein the label on the test strip is visible to the second transparentwindow of the cover.
 8. The device according to claim 3 wherein the drugof abuse is from the group consisting of methamphetamine,opiates/morphine, marijuana/tetrahydrocannabinol, amphetamine,cocaine/benzoylecgonine, methadone; PCP, habituate, trichloroacetic acidand benzodaizepine.